Inhibitory antibodies to factor Vlll develop in approximately 30% of patients with severe and moderately severe hemophilia A in response to infusions of factor Vlll. Inhibitor development is associated with a lower quality of life and an increased economic burden. Consequently, inhibitor development cunrently is considered the most significant complication of the management of in hemophilia A. Additionally, factor Vlll inhibitors can occur in nonhemophiliacs, producing an autoimmune condition called acquired hemophilia A. Recent developments have made it possible to ask increasingly focused questions regarding the pathogenesis of factor Vlll inhibitor development and to test novel diagnostic and therapeutic approaches to the problem. There are four Specific Aims in this project. In Aim 1, novel immunodominant epitopes recognized by anti-factor Vlll antibodies produced in a murine hemophilia A immunogenicity model will be identified and characterized. This will be accomplished using several methods, including domain-specific antibody mapping, homolog and site-directed mutagenesis of factor Vlll, and functional analysis of anti-factor Vlll antibodies. Additionally, the pathogenicity of anti-factor Vlll antibodies will be evaluated in hemophilia A mice. In Aim 2, mechanisms of antigen presentation during the immune response to factor Vlll in the murine hemophilia A model will be determined. Factor Vlll is an extremely potent immunogen in hemophilia A mice under conditions in which most proteins are not immunogenic. We will test the hypothesis that antigen presentation by dendritic cells and/or macrophages is more efficient in the case of factor Vlll compared with other proteins. Additionally, we will attempt to identify the underlying mechanism for this phenomenon by identifying receptors that are involved in presentation of factor Vlll. The role von Willebrand factor (VWF) plays in the immune response to factor Vlll is controversial. Our experiments will include determination of whether the immune response to factor Vlll depends on its association with VWF. In Aim 3, the immunogenicity of factor Vlll will be decreased by antigen-specific deletion of naive and memory factor B-cells.